The debilitating human parasitic disease leishmaniasis, caused by the parasitic protozoan Leishmania, is endemic in the tropical regions of America, Africa, and the Indian sub-continent, and the sub-tropics of south-west Asia and the Mediterranean. The total number of infected people in the world is estimated to be 12 millions, and 350 millions live in areas where the disease may be caught. The appearance of cases of visceral leishmaniasis in individuals infected with HIV but without a history of leishmaniasis, suggests that Leishmania may survive asymptotically in immunologically competent hosts. HIV destroys immunological competence and permits parasite multiplication. Thus, HIV infection in endemic areas of leishmaniasis poses additional threats to human health. Identification and understanding of the molecules responsible for parasite virulence is essential for development of rational chemotherapy and vaccine against the parasite infection. Several Leishmania molecules have been suggested as potential virulent factors by virtue of their high abundance on the parasite cell surface, correlation of their presence with the infectivity or their differential presence in the infective form of the parasite. Individual or collective contributions of those molecules towards the establishment and maintenance of Leishmania infection in mammalian macrophages are far from clear. The working hypothesis on which this proposal is made is that the parasites induce or suppress the expression of certain genes as required for their infectivity to mammalian system. Those genes could be the genes for the potential virulent factors or could be the genes for the enzymes that catalyze the biosynthesis of the virulent factors. The specific aims of this proposal include (i) cloning and characterization of the genes that are induced (and/or repressed) in infective variants (virulent clones of promastigotes and amastigotes) of Leishmania mexicana amazonensis employing the subtractive cloning and other techniques; and (ii) testing the efficacy of the potential cloned genes, individually or in combination, to transform avirulent cells to virulent cells. Successful completion of this project and future continued studies aimed at understanding the regulation of the expression of the virulent genes of Leishmania will not only add to better understanding of the cell biology of Leishmania-macrophage interactions but also yield information leading to the development of rational antileishmanial chemotherapy . Identification and characterization of virulent genes will also help precise vaccine development strategies by selective depletion of one or all of those genes from Leishmania and using the resultant avirulent live parasites as vaccine.